人膝骨关节炎血清生物学标志物的筛选

目的: 比较膝骨关节炎患者与正常人血清蛋白质组的差异,筛选其能用于骨关节炎诊断、治疗或发病机制研究的血清生物学标志物。方法: 利用双向荧光差异凝胶电泳技术比较膝骨关节炎患者血清(4例)与健康志愿者血清(4例)蛋白图谱的差异,使用基质辅助激光解吸电离-飞行时间质谱及生物信息学相关技术对获得的差异蛋白进行鉴定和初步分析,并用Western blotting进一步验证所鉴定的蛋白。结果: 成功建立了骨关节炎血清差异蛋白质组学的研究方法;通过质谱分析初步鉴定出8个差异蛋白,其中5个蛋白在膝骨关节炎组表达上调,3个蛋白在膝骨关节炎组表达下调。Western blotting进一步验证得到α₂-巨球蛋白在膝骨关节炎组表达上调,与双向荧光胶内差异凝胶电泳技术联合质谱分析的结果一致。结论: 膝骨关节炎患者血清与正常人血清蛋白表达存在明显差异,其中α₂-巨球蛋白可作为骨关节炎潜在的疾病相关生物学标志物进一步研究,为骨关节炎的诊断、治疗和发病机制的研究提供新的线索和实验基础。     

英夫利西单抗治疗克罗恩病黏膜愈合的血清差异蛋白质表达研究

目的:寻找与英夫利西单抗(infliximab,IFX)治疗克罗恩病(Crohn’s disease,CD)黏膜愈合(mucosal healing,MH)相关的血清蛋白质生物标志物。方法:采集7例经IFX治疗获得MH的CD患者治疗前(0周,A组)和治疗后(14周,B组)的血清,以及7例未获得MH的CD患者(0周为C组,14周为D组)的血清。采用荧光标记双向差异凝胶电泳的方法,比较A组与B组之间、C组与D组之间、A组与C组之间以及B组与D组之间的蛋白质组学差异,并对差异表达的蛋白质斑点进行基质辅助激光解吸/电离飞行时间串联质谱初步鉴定和生物信息学分析。结果:(1)A组与B组、C组与D组、A组与C组以及B组与D组之间比较分别存在36、3、10和31个差异表达的蛋白质斑点,共计存在44个显著差异表达的蛋白质斑点。(2)上述各组之间的差异斑点分别初步鉴定出17、2、2和15种蛋白质,共计存在19种差异表达的蛋白质,包括载脂蛋白E、载脂蛋白A-I、补体因子H等。(3)基于STRING数据库绘制了蛋白质网状功能图。结论:IFX治疗获得MH的CD患者治疗前后的血清蛋白质表达谱存在差异,获得MH和未获得MH的患者血清表达谱亦存在差异,其中19种蛋白质有可能成为预测CD患者使用IFX获得MH的生物标志物。     

Proteomic analysis of sputum in patients with active pulmonary tuberculosis

The protein composition of sputum most faithfully reflects the state of the lungs. The aim of this study was to determine whether relative qualitative and quantitative differences in protein expression of sputum could be related to active pulmonary tuberculosis. Sputum samples were collected from 65 patients with active pulmonary tuberculosis and 38 healthy controls. Comprehensive proteomic approaches were used to profile the proteome changes of host sputum in response to Mycobacterium tuberculosis infection using two-dimensional electrophoresis in combination with matrix-assisted laser desorption ionization time-of-flight/time-of-flight mass spectrometry. Mascot software was used to identify proteins from protein databases. Enzyme-linked immunosorbent assay was used to confirm the proteomic results. A total of 62 differentially expressed proteins were identified, among which, 15 proteins were up-regulated and 47 proteins were down-regulated in the tuberculosis sputum compared with the controls. Bacterial protein UqhC was the most increased protein, whereas serum albumin was the most decreased protein in the tuberculosis sputum compared with the controls. The enzyme-linked immunosorbent assay analysis was consistent with proteomic data. Bioinformatics analysis suggested that multiple host cell pathways were involved in the tuberculosis infection processes, including acute phase response, signal transduction, cytoskeleton structure, immune response and so on. In all, for the first time, our results revealed that a number of proteins were differentially expressed during active pulmonary tuberculosis infection. These data will provide valuable clues for further investigation of tuberculosis pathogenesis and biomarkers for detection of active pulmonary tuberculosis infection.     

应用TMT结合质谱技术筛选高原红细胞增多症患者血浆差异表达蛋白

目的:应用串联质谱标签(TMT)结合质谱技术比较高原红细胞增多症(HAPC)患者和健康对照者血浆中的差异表达蛋白。方法:收集HAPC患者血浆4例,与之相匹配的健康对照组血浆5例,组内等量混合后利用多重免疫亲和层析柱去除14种高丰度蛋白,TMT试剂标记后样本进行液相色谱分离,采用HPLC-MS/MS质谱鉴定及相对定量;获取HAPC患者和健康对照血浆各20例,ELISA方法验证部分筛选的差异蛋白。结果:共鉴定和定量了1 094种蛋白,差异表达的蛋白有249种,其中HAPC组较健康对照组表达量≥1.5倍的上调蛋白质有162种,表达量≤67%的下调蛋白质有87种;C反应蛋白和血清淀粉样蛋白A在HAPC组中较健康对照组表达量均上调(P0.05)。结论:筛选出多种与炎症相关的差异表达蛋白,为深入研究HAPC发生机制奠定了良好基础。     

MALDI-TOF MS combined with magnetic beads for detecting serum protein biomarkers and establishment of boosting decision tree model for diagnosis of colorectal cancer

The aim of present study is to study the serum protein fingerprint of patients with colorectal cancer (CRC) and to screen protein molecules that are closely related to colorectal cancer during the onset and progression of the disease with Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Serum samples from 144 patients with CRC and 120 healthy volunteers were adopted in present study. Weak cation exchange (WCX) magnetic beads and PBSII-C protein chips reader (Ciphergen Biosystems Ins.) were used. The protein fingerprint expression of all the Serum samples and the resulted profiles between cancer and normal groups were analyzed with Biomarker Wizard system. Several proteomic peaks were detected and four potential biomarkers with different expression profiles were identified with their relative molecular weights of 2870.7 Da, 3084 Da, 9180.5 Da, and 13748.8 Da, respectively. Among the four proteins, two proteins with m/z 2870.7 and 3084 were down-regulated, and the other two with m/z 9180.5 and 13748.8 were up-regulated in serum samples from CRC patients. The present diagnostic model could distinguish CRC from healthy controls with the sensitivity of 92.85% and the specificity of 91.25%. Blind test data indicated a sensitivity of 86.95% and a specificity of 85%. The result suggested that MALDI technology could be used to screen critical proteins with differential expression in the serum of CRC patients. These differentially regulated proteins were considered as potential biomarkers for the patients with CRC in the serum and of the potential value for further investigation.     

Noticeable Decreased Expression of Tenascin-X in Calcific Aortic Valves

Calcification of aortic valves results in valvular aortic stenosis and is becoming a common valvular condition in elderly populations. An understanding of the molecular mechanisms of this valve lesion is important for revealing potential biomarkers associated with the development and progression of this disease. In order to identify proteins that are differentially expressed in calcific aortic valves (CAVs) compared with those in adjacent normal valvular tissues, comprehensive analysis of differentially expressed proteins in the tissues was done by a quantitative proteomic approach with isobaric tag for absolute and relative quantitation labeling followed by nanoliquid chromatography matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry. The proteomic analysis revealed 105 proteins differentially expressed in CAVs in contrast to adjacent normal valvular tissues with high confidence. Significantly increased expression (>_1.3-fold) was found in 34 proteins, whereas decreased expression (<0.77-fold) was found in 39 proteins in CAVs. Among them, α-2-HS-glycoprotein showed the greatest increase in expression (6.54-fold) and tenascin-X showed the greatest decrease in expression (0.37-fold). Numerous extracellular matrix proteins such as collagens were identified as proteins with significantly decreased expression. Panther pathway analysis showed that some of the identified proteins were linked to blood coagulation and integrin signaling pathways. Cluster analysis of the 105 proteins differentially expressed in CAVs based on the expression pattern revealed that tenascin-X was clustered with proteins controlling collagen structure and function, especially collagen fibrillogenesis, such as decorin and fibromodulin. We confirmed decreased levels of these proteins in CAVs by Western blot analyses. These results indicated that massive destruction of the extracellular matrix occurs in CAVs.     

Comparative analysis of serum proteomes: discovery of proteins associated with osteonecrosis of the femoral head

No means exist to evaluate the activity status, turnover, and prognosis of idiopathic osteonecrosis of the femoral head (IONFH) except for X-ray evidence of segmental collapse as a very good marker for prognosis. Moreover, the only current method for diagnosis of this disease is through physical examination and diagnostic imaging results, and no serum biochemical markers exist. A comparative analysis of serum proteomes was performed to discover proteins associated with osteonecrosis of the femoral head. Two-dimensional electrophoresis (2-DE) patterns of human sera from 10 patients with IONFH and 10 normal subjects were analyzed. The differentially expressed spots were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and 7 proteins were found. The expression levels of tissue-type plasminogen activator (t-PA), bone-carboxyglutamate protein (BGP), c-sis, and an unknown protein were downregulated in the sera of patients with IONFH, whereas the other 3 proteins, including plasminogen activator inhibitor type 1 (PAI-1), crosslaps, and anti-p53 antibody, were upregulated. To examine their applicability as diagnostic markers, levels of the 6 identified proteins in serum were validated from patients with IONFH, osteoarthritis (OA), rheumatoid arthritis (RA), and fracture using the enzyme-linked immunosorbent assay (ELISA) method. It was found that only serum levels of t-PA, PAI-1, crosslaps, and anti-p53 antibody in patients with IONFH were always significantly different from those in patients with OA, RA, and fracture. These results suggest that serum levels of t-PA, PAI-1, crosslaps, and anti-p53 antibody could be used as noninvasive diagnostic biomarkers for IONFH.     

Proteomic profiling of mature CD10+ B-cell lymphomas

Proteomic profiling with protein-chip technology has been used successfully to discover biomarkers with potential clinical usefulness in several cancer types. Little proteomic study has been done in B-cell lymphomas. We determined whether the expression of a set of proteins by protein-chip technology coupled with new informatics tools could be used to build a model to molecularly classify B-cell lymphoma subgroups. We used surface-enhanced laser desorption/ionization time-of-flight mass spectrometry to analyze 18 CD10+ B-cell lymphomas, including 6 grade 1 (G1) follicular lymphomas (FLs), 7 grade 3 (G3) FLs, and 5 Burkitt lymphomas. We used 7 reactive follicular hyperplasia cases as a control group. By using SAX2 ProteinChip arrays (Ciphergen Biosystems, Fremont, CA), we found a unique protein expression profile for each type of lesion. Two-way hierarchical clustering analysis of these protein expression profiles differentiated reactive follicular hyperplasia, FL, and Burkitt lymphoma, with 5 major clusters of differentially expressed protein peaks. In addition, we identified histone H4 as a potential differentially expressed protein marker that seems to distinguish G1 from G3 FL. To our knowledge, this is the first proteomic study using protein-chip technology for molecular classification of B-cell lymphoma subtypes with clinical samples.     

帕金森病相关血清蛋白质标志物的筛选及鉴定

 目的：探讨并初步鉴定帕金森病患者血清中相关蛋白质作为特异标志物的可能性。方法：应用表面增强激光解吸电离飞行时间质谱(SELDI-TOF-MS)结合纳米磁珠技术检测44例帕金森病和60例健康对照的血清标本,应用生物信息学方法筛选差异蛋白峰,经高效液相色谱（HPLC）分离出差异蛋白,酶解后进行液质联用串联质谱（LC-MS/MS）分析,利用Xcalibur的程序组件BioWorks 3.2完成蛋白质序列数据库鉴定分析。结果：经SELDI-TOF-MS结合纳米磁珠技术筛选出质荷比m/z位于8 937的蛋白质在帕金森病中高表达(帕金森病组表达强度为27.47±16.58,正常组表达强度为5.01±3.47),有显著差异(P<0.01);6 636和8 697的蛋白质在帕金森病中低表达(帕金森病组表达强度为5.43±2.66和20.22±9.57,正常组表达强度为18.85±7.56和51.13±26.22), 有显著差异(P<0.01)。联合上述3种潜在蛋白质标志物,可区分帕金森病组和对照组,其中帕金森病患者检出率为90.0%(27/30),健康者检出率为92.5%（37/40）。对m/z为6 636、8 697和8 937的标志物进行鉴定,结果分别为载脂蛋白C-I、载脂蛋白 C-III 和补体成分3a。结论：鉴定出的载脂蛋白 C-I、载脂蛋白C-III和补体成分3a在帕金森病的诊断中具有一定价值,值得进一步研究和探讨。     

Serum proteomics with SELDI-TOF-MS in congenital human cytomegalovirus hepatitis

Human cytomegalovirus (HCMV) is a widespread pathogen, the most common congenital viral infection, and the leading cause of infant hepatitis syndrome. In this study, serum samples were collected from 20 HCMV-infected infants with hepatitis and 25 controls. Of the 25 infants in the control group, 5 were infected with HCMV but without hepatitis, 10 had hepatitis but no HCMV infection, and 10 were healthy. Proteomic expression in the serum was detected by WCX2 chips and surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS), to identify serum protein biomarkers in infants with hepatitis syndrome resulting from HCMV. Fifteen protein peaks were distinctly different among the four groups in the mass range from 2,000 to 20,000 Da. Of these 15 peaks, 4 at 4,349.8, 5,808.7, 7,935.6, and 8,885.9 Da were significantly different between the congenital HCMV-infected infants with hepatitis and the controls. Five peaks were distinctly up-regulated in the infants with HCMV infection (3,266.8, 5,638.5, 5,909.1, 7,771.4, and 15,835.6 Da) compared to those without HCMV infection. Two proteins at 4,600.1 and 5,704.3 were up-regulated in infants with HMCV infection but no hepatitis. Four protein peaks were markedly different (7,567.0, 13,744.8, 15,100.7, and 15,915.0 Da) between the infants with hepatitis and the other controls. Comparison of the differentially expressed proteins' properties with those available on an international database suggest that specific serum proteins such as the augmenter of liver regeneration, pre-albumin, and haptoglobin closely related to liver function, and cytokines such as beta-defensins 31 and 8, and macrophage-derived chemokine, among others, are involved in HMCV infection and the pathogenesis of HMCV-induced hepatitis in infants.     

侵袭性肺曲霉病小鼠肺组织差异表达蛋白的初步分析

目的初步分析侵袭性肺曲霉病(IPA)小鼠肺组织蛋白质的表达变化。方法运用双向电泳(2-DE)技术对正常组、正常感染组和IPA组小鼠肺组织总蛋白质进行分离,获得蛋白质表达谱;应用基质辅助激光解吸电离串联飞行时间质谱(MALDI-TOF/TOF)结合生物信息学进行蛋白质鉴定。结果初步鉴定出4个蛋白质点分别为过氧化物还原酶(peroxiredoxin-6,Prx6)、黄素还原酶(flavin reductase,FR)、Rho因子鸟苷酸解离抑制蛋白2(rho GDP-dissociation inhibitor 2,RhoGDI2)、肌球蛋白轻链2(myosin regulatory light chain 2,MLC2),它们与氧化还原平衡、信号转导通路、细胞骨架等功能有关。结论鉴定的差异表达蛋白可能与IPA的发病机制有关。     

不同临床类型原发鼻咽癌组织差异表达蛋白

 目的 寻找上行型、下行型和混合型不同临床类型鼻咽癌原发病瘤灶组织中的差异表达蛋白。方法 采用双向凝胶电泳对不同临床类型鼻咽癌患者的鼻咽癌组织和正常人鼻咽组织总蛋白进行分离,对比找出差异表达蛋白质斑点,结合基质辅助激光解吸电离-飞行时间质谱分析技术和数据库信息检索鉴定差异蛋白点。结果 与正常组比较,混合型鼻咽癌组出现31个差异蛋白质点,其中18个表达上调,13个表达下调;从混合型鼻咽癌组选择10个差异蛋白质斑点进行质谱鉴定,成功鉴定出5个,其中表达上调的有热休克固有蛋白、α1抗胰蛋白酶和巨噬细胞帽蛋白;表达下调的有S100A9蛋白和 蛋白4.1。与下行型鼻咽癌组比较,上行型鼻咽癌组出现31个差异表达蛋白点,其中15个表达上调、16个表达下调;从上行型组选择10个差异蛋白质斑点进行质谱鉴定,成功鉴定出6个,其中表达上调的有线粒体顺乌头酸酶、乙醇脱氢酶、Rab GDP解离抑制因子β;表达下调的有NM23-H1、核氯离子通道蛋白27和乙醛脱氢酶X。结论 不同临床类型鼻咽癌患者肿瘤转移相关蛋白、能量代谢蛋白、解毒蛋白与抗肿瘤转移蛋白表达水平存在差异,这些蛋白可作为临床分型标记物。     

Differential expressions of plasma proteins in systemic lupus erythematosus patients identified by proteomic analysis

IntroductionSystemic lupus erythematosus (SLE) is a chronic and complex autoimmune disease with a wide range of clinical manifestations that affects multiple organs and tissues. Therefore the differential expression of proteins in the serum/plasma have potential clinical applications when treating SLE.MethodsWe have compared the plasma/serum protein expression patterns of nineteen active SLE patients with those of twelve age-matched and gender-matched healthy controls by proteomic analysis. To investigate the differentially expressed proteins among SLE and controls, a 2-dimensional gel electrophoresis coupled with high-resolution liquid chromatography tandem mass spectrometry was performed. To further understand the molecular and biological functions of the identified proteins, PANTHER and Gene Ontology (GO) analyses were employed.ResultsA total of 14 significantly expressed (p < 0.05, p < 0.01) proteins were identified, and of these nine were up-regulated and five down-regulated in the SLE patients. The functional enrichment analysis assigned the majority of the identified proteins including alpha 2 macroglobulin, complement C4, complement factor H, fibrinogen beta chain, and alpha-1-antitrypsin were part of the complement/coagulation cascade, which is an important pathway that plays a crucial role in SLE pathogenesis. In addition to these proteins the differential expressions of ceruloplasmin, transthyretin, and haptoglobin play a potential role in the renal system abnormalities of SLE.ConclusionTherefore, the identified differentially expressed proteins are relevant to SLE patient's cohort. Most importantly the up-regulated proteins might be the potential candidates for renal system involvement in SLE disease pathogenesis. In order to confirm the diagnostic/therapeutic potential of the identified proteins, future validation studies are required.     

Evaluation of serum diagnosis of pancreatic cancer by using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry

Proteomic methods have been widely used in disease marker discovery research. The aim of this study was to discover potential biomarkers for pancreatic cancer (PCa) using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS). Crude serum samples from 132?patients with PCa and 67?healthy controls (HCs) were analyzed in duplicate using SELDI. Support vector machine (SVM) analysis of the spectra was used to generate a predictive algorithm based on proteins that were maximally differentially expressed between patients with PCa and the HCs in the training cohort. This algorithm was tested using leave-one-out cross-validation in the test cohort. From the 4?significant peaks in the training cohort, a classifier for separating patients with PCa from HCs was developed. The classifier was challenged with all samples achieving 96.67% sensitivity and 100% specificity in the training cohort and 93.1% sensitivity and 78.57% specificity in the test cohort. Additionally, the classifier correctly classified 12/12 stage?Ia and 13/16 stage?IIa PCa cases. The combination of the SELDI panel and CA19-9 was superior to CA19-9 alone in distinguishing individuals with PCa from the healthy subject group. These results suggest that high-throughput proteomic profiling has the capacity to provide new biomarkers for the early detection and diagnosis of PCa.     

Specific alterations in plasma proteins during depressed,manic, and euthymic states of bipolar disorder

Bipolar disorder (BD) is a common psychiatric mood disorder affecting more than 1-2% of the general population of different European countries. Unfortunately, there is no objective laboratory-based test to aid BD diagnosis or monitor its progression, and little is known about the molecular basis of BD. Here, we performed a comparative proteomic study to identify differentially expressed plasma proteins in various BD mood states (depressed BD, manic BD, and euthymic BD) relative to healthy controls. A total of 10 euthymic BD, 20 depressed BD, 15 manic BD, and 20 demographically matched healthy control subjects were recruited. Seven high-abundance proteins were immunodepleted in plasma samples from the 4 experimental groups, which were then subjected to proteome-wide expression profiling by two-dimensional electrophoresis and matrix-assisted laser desorption/ionization-time-of-flight/time-of-flight tandem mass spectrometry. Proteomic results were validated by immunoblotting and bioinformatically analyzed using MetaCore. From a total of 32 proteins identified with 1.5-fold changes in expression compared with healthy controls, 16 proteins were perturbed in BD independent of mood state, while 16 proteins were specifically associated with particular BD mood states. Two mood-independent differential proteins, apolipoprotein (Apo) A1 and Apo L1, suggest that BD pathophysiology may be associated with early perturbations in lipid metabolism. Moreover, down-regulation of one mood-dependent protein, carbonic anhydrase 1 (CA-1), suggests it may be involved in the pathophysiology of depressive episodes in BD. Thus, BD pathophysiology may be associated with early perturbations in lipid metabolism that are independent of mood state, while CA-1 may be involved in the pathophysiology of depressive episodes.     

Sputum proteomic analysis for distinguishing between pulmonary tuberculosis and non-tuberculosis using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS): preliminary results

ObjectivesThe aim was to evaluate the feasibility and diagnostic contribution of protein profiling using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) applied to sputum to diagnose pulmonary tuberculosis.MethodsSputum samples collected from patients suspected of having pulmonary tuberculosis were analysed using MALDI-TOF MS. Using the differentially expressed protein peaks, we compared three groups of patients, including those with confirmed pulmonary tuberculosis (PTB), those without tuberculosis but with a lower respiratory tract infection (non-TB LRTI) and those without tuberculosis and without an LRTI (non-TB controls).ResultsA total of 102 patients included 35 PTB, 36 non-TB LRTI and 31 non-TB controls. The model differentiated between the PTB patients and the non-TB controls using the 25 most differentially expressed protein peaks, with a sensitivity of 97%, 95% CI 85–100%, and a specificity of 77%, 95% CI 59–90%. The model distinguished the PTB patients from the non-TB LRTI patients using the ten most differentially expressed protein peaks, with a sensitivity of 80%, 95% CI 63–92%, and a specificity of 89%, 95% CI 74–97%. We observed that the negative predictive value of MALDI-TOF MS sputum analysis was higher (96%, 95% CI 80–100%) than that of direct sputum microscopic examination and sputum culture (78%, 95% CI 62–89%) for non-TB controls. When MALDI-TOF MS sputum analysis and direct microscopic examination were combined, the negative predictive value reached 94%, 95% CI 80–99%, for non-TB LRTI patients.DiscussionThese results suggest that MALDI-TOF MS sputum analysis coupled with microscopic examination could be used as a screening tool for diagnosing pulmonary TB.     

Serum microRNAs as novel biomarkers for primary sclerosing cholangitis and cholangiocarcinoma

The diagnosis of primary sclerosing cholangitis (PSC) is difficult due to the lack of sensitive and specific biomarkers, as is the early diagnosis of cholangiocarcinoma (CC), a complication of PSC. The aim of this study was to identify specific serum miRNAs as diagnostic biomarkers for PSC and CC. The levels of 667 miRNAs were evaluated in 90 human serum samples (30 PSC, 30 CC and 30 control subjects) to identify disease‐associated candidate miRNAs (discovery phase). The deregulated miRNAs were validated in an independent cohort of 140 samples [40 PSC, 40 CC, 20 primary biliary cirrhosis (PBC) and 40 controls]. Receiver operating characteristic (ROC) curves were established and only miRNAs with an area under the curve (AUC) > 0·70 were considered useful as biomarkers. In the discovery phase we identified the following: 21 miRNAs expressed differentially in PSC, 33 in CC and 26 in both in comparison to control subjects as well as 24 miRNAs expressed differentially between PSC and CC. After the validation phase, miR‐200c was found to be expressed differentially in PSC versus controls, whereas miR‐483‐5p and miR‐194 showed deregulated expression in CC compared with controls. We also demonstrate a difference in the expression of miR‐222 and miR‐483‐5p in CC versus PSC. Combination of these specific miRNAs further improved the specificity and accuracy of diagnosis. This study provides a basis for the use of miRNAs as biomarkers for the diagnosis of PSC and CC.     

慢性重型肝炎患者血清中的低分子量蛋白质研究

目的：应用蛋白质芯片技术平台研究慢性重型肝炎血清中的低分子量蛋白质。方法:利用表面增强激光解吸离子化飞行时间串联质谱技术，检测28例慢性重型肝炎患者和22例健康对照组血清中相对分子质量位于1 kD-10 kD之间的蛋白质指纹图谱，筛选两组图谱的差异峰，结合主成分分析方法区分健康对照组和重型肝炎患者；对慢性重型肝炎组和健康对照组的血清样本进行超滤处理，研究超滤前后血清蛋白质指纹图谱的变化情况。结果:比较慢性重型肝炎组与健康对照组的蛋白质指纹图谱，有39个峰强度在两组间的P<1×10-6。用主成分分析方法对得到的数据进行处理，在得到的投影结果图上，健康对照组与慢性重型肝炎组的划分边界非常清楚。对比血清超滤前后的蛋白质指纹图谱，发现在经过截留率为30 kD的超滤膜超滤后，超滤液中相对分子质量位于1 kD～10 kD的峰基本消失。结论:慢性重型肝炎患者的血清中存在具有明显差异的低分子量蛋白质。     

Proteomic analysis of calcified abdominal and thoracic aortic aneurysms

Aortic aneurysm is a complex multifactorial disease with genetic and environmental risk factors. It is often accompanied by aortic calcification. Here, to uncover proteins that are significantly changed in calcified abdominal aortic aneurysms (CAAs) and calcified thoracic aortic aneurysms (CTAs) compared with those in adjacent normal aorta tissues, comprehensive analysis of differentially expressed proteins in their tissues was performed by a quantitative proteomic approach with iTRAQ labeling in combination with nanoLC-MALDI-TOF/TOF-MS/MS followed by ProteinPilot analysis. The proteomic analysis revealed 138 and 134 proteins differentially expressed in CAAs and CTAs in contrast to neighboring normal aorta tissues with high confidence, respectively. Significantly increased expression (≥1.3-fold) was found in 41 and 28 proteins, whereas decreased expression (<0.77-fold) was found in 4 and 60 proteins in CAAs and CTAs, respectively. Among them, we identified already known proteins involved in aneurysm formation and vascular calcification, such as type I and III collagen, matrix Gla protein, and α-2-HS-glycoprotein in CAAs and fibrinogen α, β and γ chains and α-2-HS-glycoprotein in CTAs with increased expression and mimecan in CAAs and fibulin-5 in CTAs with decreased expression. Based on the Panther pathway and Genesis clustering analyses, some of the proteins could be linked to corresponding biochemical pathways, such as the integrin signaling pathway with increased expression in CAAs, the blood coagulation pathway with increased expression in CTAs, and the inflammation mediated by chemokine and cytokine signaling pathway and the glycolysis pathway with decreased expression in CTAs. Interestingly, it was found by clustering analysis that samples from CAAs of patients with both CAAs and CTAs were clustered outside the samples of patients with CAAs and were clustered with samples of patients with CTAs. Our results provide a comprehensive patient-based proteomic analysis for the identification of potential biomarkers for CAAs and CTAs.     

Elevated expression of complement C3 protein in chemically induced hepatotumorogenesis in Wistar rats: A correlative proteomics and histopathological study

Liver cancer remains the leading cause of cancer-related mortality worldwide. Early detection of liver cancer is problematic due to the lack of a marker with high diagnosis sensitivity and specificity. The present study was designed to determine the differently expressed proteins at early stage in the serum of animals with liver cancer vis-à-vis controls and figure out the function of the proteins. One-dimensional electrophoresis (1D), two-dimensional electrophoresis (2DE) and liquid chromatography mass spectrometry (LC–MS/MS) were used to screen the serum proteins of liver cancer induced in animals by diethyl nitrosamine (DEN) + 2-acetyl amino fluorine (2-AAF). From optimized 2DE image and computer assisted PD Quest analysis were found to be differentially expressed spots when the serum from normal and treated animals were compared. Among these, one spot was selected whose expression level was higher in DEN + 2-AAF treated animal sera than in adjacent normal animal sera. The target spot was excised from the 2D gel of liver cancer sera and the peptide mass fingerprinting as obtained LC–MS/MS analysis after digesting the chosen protein spot. This was identified to be complement C3 protein. The changes in complement C3 expression level were validated by Western blot analysis. We reported that the changes in complement C3 concentration start at very early stage of tumorogenesis. The fully grown tumors were developed at 120 days and hepatotumorogenesis was confirmed by histopathological examination. This protein may therefore represent a powerful tool in search for candidate biomarkers for HCC.